Covalent modification of cellular substrates with methyl groups has been implicated in the pathology of cancer and other diseases. (Gloria, L. et al. (1996) Cancer 78:2300-2306.) Cytosine hypermethylation of eukaryotic DNA prevents transcriptional activation. (Turker, M. S. and Bestor, T. H. (1997) Mutat. Res. 386:119-130.) N.sup.6 -methyladenosine is found at internal positions of mRNA in higher eukaryotes. (Bokar, J. A. et al. (1994) J. Biol. Chem. 269:17697-17704.) Hypermethylated viral DNA is transcribed at higher rates than hypo- or hemimethylated DNA in infected cells. (Willis, D. B. et al. (1989) Cell. Biophys. 15:97-111.)
Propagation of nerve impulses, modulation of cell proliferation and differentiation, induction of the immune response, and tissue homeostasis may involve neurotransmitter metabolism. (Weiss, B. (1991) Neurotoxicology 12:379-386; Collins, S. M. et al. (1992) Ann. N.Y. Acad. Sci. 664:415-424; and Brown, J. K. and Imam, H. (1991) J. Inherit. Metab. Dis. 14:436-458.) In tissue, synthesis and rates of degradation that regulate the activity of neurotransmitters are dependent upon enzyme and cofactor levels. (Brown, J. K. and Imam, H. supra.) Many pathways of small molecule degradation, such as those of neurotransmitters, require methyltransferase activity. (Kagan, R. M. and Clarke, S. (1994) Arch. Biochem. Biophys. 310:417-427.) For example, degradation of the catecholamines epinephrine or norepinephrine, requires catechol-O-methyltransferase, and N-acetyl-5-hydroxytryptamine is converted to melatonin by hydroxyindole-O-methyltransferase in the pineal gland. Both catechol-O-methyltransferase and hydroxyindole methyltransferase genes contain alternative initiation codons. (Rodriguez, I. R. et al. (1994) J. Biol. Chem. 269:31969-31977; and Tenhunen, J. et al. (1994) Eur. J. Biochem. 223:1049-1059.)
S-adenosylmethionine (AdoMet) is an important source of methyl groups for methylation reactions in the cell. (Bottiglieri, T. and Hyland, K. (1994) Acta Neurol. Scand. Suppl. 154:19-26.) Methyltransferase activity catalyzes the transfer of methyl groups from AdoMet to acceptor molecules such as phosphotidylethanolamine or the polynucleotide 5' cap of viral mRNA. (Montgomery, J. A. et al. (1982) J. Med. Chem. 25:626-629.)
Members of the protein and small molecule S-adenosylmethionine methyltransferase family (AdoMet-MT), utilize AdoMet as a substrate or product and harbor three common consensus sequence motifs. (Kagan and Clarke, supra.) Motifs I and II are characteristically spaced between 34 and 90 (mode 52, mean 57.+-.13) amino acid residues apart; motifs II and III are spaced between 12 and 38 (mode 22, mean 22.+-.5) residues apart. Motif I comprises part of the AdoMet binding pocket; motif II may also be involved in binding AdoMet; the role of motif III is uncertain. The main exceptions to the spacing rule are the RNA methyltransferases and a number of the porphyrin precursor methyltransferases. It has been suggested that these heterogeneic motifs may be of use in predicting methyltransferases and related enzymes from open reading frames generated genomic sequencing projects. (Kagan and Clarke, supra.)
Messenger RNA N.sup.6 -adenosine methyltransferase holoenzyme has been partially purified from HeLa cell nuclear extract to yield three subunits, an 875 kDa ssDNA-agarose binding protein, a 70 kDa AdoMet-binding protein, and an approximately 30 kDa component with unknown function. The three components are absolutely required for RNA m.sup.6 A-methylation activity. (Bokar, J. A., supra.)
In many tissues, including brain, gut, bone marrow, liver, and kidney, serine hydroxymethyltransferase converts serine to glycine by transferring the hydroxymethyl side chain group of serine to the methyl acceptor, tetrahydrofolate. The product of this reaction is N.sup.5,N.sup.10 -methylenetetrahydrofolate and water. N.sup.5,N.sup.10 -methylenetetrahydrofolate is a substrate in de novo purine nucleotide synthesis and pyrimidine nucleotide synthesis, in conversion of homocysteine to methionine, and in methylation of tRNA, during tissue growth and cell proliferation.
The genes encoding many of the growth-associated methyltransferases have not yet been identified or isolated. In their roles as a rate-limiting step in methyltransferase reactions, AdoMet-MTs have been identified as a target for psychiatric, antiviral, anticancer and anti-inflammatory drug design. (Bottiglieri, T. and Hyland, K., supra; Gloria, L. et al., supra.) Sequence-specific methylation inhibits the activity of the Epstein-Barr virus LMP1 and BCR2 enhancer-promoter regions. (Minarovits, J. et al. (1994) Virology 200:661-667.) 2'-5'-linked oligo (adenylic acid) nucleoside analogues synthesized by interferon-treated mouse L cells act as antiviral agents. (Goswami, B. B. et al, (1982) J. Biol. Chem. 257:6867-6870.) Adenine analogue inhibitors of AdoMet-MT decreased nucleic acid methylation and proliferation of leukemia L1210 cells. (Kramer, D. L. et al. (1990) Cancer Res. 50:3838-3842.)
The use of experimental neuroactive drugs has shown that inactivation of neurotransmitters is absolutely essential for the correct functioning of the nervous system. (Avery, L. and Horvitz, H. R. (1990) J. Ex. Zool. 253:263-270.) Epigenetic or genetic defects in neurotransmitter metabolic pathways can result in a spectrum of disease states in different tissues including Parkinson disease and inherited myoclonus. (McCance, K. L. and Huether, S. E. (1994) Pathophysiology, Mosby-Year Book, Inc., St. Louis, Mo. pp. 402-404; and Gundlach, A. L. (1990) FASEB J. 4:2761-2766.)
The discovery of two new human growth-associated methyltransferases and the polynucleotides encoding them satisfies a need in the art by providing new compositions which are useful in the diagnosis, treatment, and prevention of neoplastic, immunological, reproductive, developmental, and vesicle trafficking disorders.